Figure 6.

Allelic mismatching of the Pik helper NLRs can be used to alleviate autoactivity from nanobody integration in Pikm-1. Five pikobodies that either respond to the FP eGFP (Pikm-1^LaG24 and Pik-1^Enhancer indicated by green text) or mCherry (Pikm-1^LaM2, Pikm-1^LaM3, and Pikm-1^LaM6, indicated by red text) were tested with the Pikm-2 or Pikp-2 helpers to assess the effect of mismatching on cell death responses in N. benthamiana. A), B) Pikm-1^LaG24 and Pikm-1^LaM6 are autoactive in the presence of Pikm-2; however, coexpression with the Pikp-2 helper does not result in autoactivity. Both pikobodies retain ability to respond to their respective FPs. C) Pikm-1^Enhancer is not autoactive in the presence of both helpers and can respond to eGFP with equal strength. D) Coexpression of Pikm-1^LaM2 with Pikp-2 alleviates autoactivity observed in the presence of Pikm-2; however, response to mCherry is substantially reduced. E) Mismatching of the helper alleles is unable to prevent autoactivity induced by the Pikm-1^LaM3 pikobody. Details of the NLR mutants used in these experiments can be found in Supplemental Table S3.